1. Field of Industrial Application
The present invention relates to a capillary having adsorbed a polymer, and a process for producing the same.
2. Prior Art
Capillary electrophoresis (hereinafter sometimes called as CE) is a generic name of electrophoresis conducted in a capillary having an inner diameter of 100 xcexcm or less. The CE has characteristic features, such as super high resolving power (number of theoretical plates: 100,000 to 10,000,000), high speed analysis, a minute amount of sample (several nl), the use of an aqueous solvent, possibility of capillary on-line detection, and the like. By utilizing these characteristic features, the CE is used in various fields, such as measurement of drugs, foods and environmental substances. Among these, analysis of proteins can be exemplified as a filed that has a possibility of further development by the CE. However, it is considered that hydrophobic silicon and a polar silanol group are present on an inner wall of a capillary, and therefore in analysis of proteins by the CE, tailing of peaks or irreversible adsorption of a solute on the wall are often found due to an ionic mutual interaction and a hydrophobic interaction between the solute and the inner wall of the capillary. As a solution thereof, a capillary having an inner wall coated with a hydrophilic substance is employed.
The capillaries having a coated inner wall known at the present time have many problems, such as lack of stability, difference among lots, an insufficient adsorption preventing function, a high cost, and the like, and a technique for solving these problems is further demanded.
The invention relates to (1) a capillary having adsorbed on an inner wall of the capillary, an ionic polymer having positive electric charge and an ionic polymer having negative electric charge alternately, (2) a capillary having adsorbed on an inner wall of the capillary, an ionic polymer having positive electric charge and an ionic polymer having negative electric charge alternately, the innermost layer being the ionic polymer having positive electric charge, (3) a capillary having adsorbed on an inner wall of the capillary, an ionic polymer having positive electric charge and an ionic polymer having negative electric charge alternately, the innermost layer being the ionic polymer having negative electric charge, (4) a process for producing a capillary characterized by making the capillary adsorb, on an inner wall of the capillary, an ionic polymer having positive electric charge and an ionic polymer having negative electric charge alternately, (5) a process for producing a capillary, in the process for producing a capillary described above, comprising the step of firstly rinsing the capillary with a strong alkali solution before making it adsorb the ionic polymer having positive electric charge.
The capillary according to the present invention means a capillary for capillary electrophoresis, a capillary for connecting in gas chromatography and mass spectrometry, and the like, and can be applied to an infusion analysis of mass spectrometry and capillary electrophoresis/mass spectrometry.
The material of the capillary is generally fused quartz glass, and the inner diameter is generally from 50xcexc to 100xcexc.
The ionic polymer having positive electric charge in the present invention means a polymer having positive electric charge that is dissolved in water or an aqueous organic solvent, such as methanol, ethanol, acetonitrile, etc., and specifically, polybrene, chitosan, diethylaminoethyldextran, polyethyleneimine, etc. can be exemplified. These substances are generally used singly, but may be used in combination. Furthermore, different substances may be used in respective layers.
The ionic polymer having negative electric charge in the present invention means a polymer having negative electric charge that is dissolved in water or an aqueous organic solvent, such as methanol, ethanol, acetonitrile, etc., and specifically, dextran sulfate, heparin, heparin sulfate, hyaluronic acid, chondroitin sulfate, keratan sulfate, polygalacturonic acid, arginic acid, teikronic acid, etc. can be exemplified. These substances are generally used singly, but may be used in combination. Furthermore, different substances may be used in respective layers.
In the capillary according to the present invention, when the ionic polymer having positive electric charge and the ionic polymer having negative electric charge are adsorbed alternately in the same number of times, the innermost layer (the layer in contact with an electrophoretic solution) is the ionic polymer having negative electric charge, and when the number of times of adsorption of the ionic polymer having positive electric charge is larger than that of the ionic polymer having negative electric charge by once, the innermost layer is the ionic polymer having positive electric charge. In general, the ionic polymer having positive electric charge and the ionic polymer having negative electric charge each are coated once alternately, or the ionic polymer having positive electric charge, the ionic polymer having negative electric charge and then the ionic polymer having positive electric charge are coated alternately in this order, but both of the layers may be coated alternately twice or more.
Upon producing the capillary according to the present invention, in the case where the ionic polymer having positive electric charge and the ionic polymer having negative electric charge each are coated once alternately, it can be produced in such a manner that the capillary is rinsed with 1N sodium hydroxide and then water, and then after rinsing with the ionic polymer having positive electric charge of a low concentration, the capillary is rinsed with the ionic polymer having positive electric charge of a high concentration, followed by immediately rinsing with the ionic polymer having negative electric charge of a high concentration.
A capillary having adsorbed two or more layers of the ionic polymers can be produced by repeating the operation described above except for the rinse with 1N sodium hydroxide and the subsequent rinse with water. The rinse with 1N sodium hydroxide may be conducted with other strong alkali solutions, such as potassium hydroxide, etc.
The concentration of the ionic polymer having positive electric charge of a low concentration, which cannot be determined unconditionally as is different depending on the species of the ionic polymer, generally means a low concentration within a concentration range in that an electrosmosis flow does not largely depend on the concentration of the ionic polymer, and is, for example, from 0.1 to 0.5%, and preferably about 0.4%, for polybrene. The ionic polymer having positive electric charge of a high concentration means a high concentration within a concentration range in that an electrosmosis flow does not largely depend on the concentration of the ionic polymer, and is, for example, from 3 to 10%, and preferably from 5 to 10%, for polybrene.
The concentration of the ionic polymer having negative electric charge of a high concentration is preferably as high concentration as possible as far as it is soluble in a solvent, but since the viscosity of the solvent is generally increased at a high concentration to make rinsing difficult, the concentration is determined on balance between them, and is generally from 1 to 5%. For example, it is about 2% for dextran sulfate and is about 3% for heparin.